| Hepatocellular carcinoma is one of the most common malignant tumor. Every year,approximately640thousand cases are newly diagnosed as HCC worldwidely, and morethan half are Chinese. In China, HCC is at the second place among the most frequentcauses for cancer-related deaths since1990s; and HBV infection is the primary etiologythat leads to liver cirrhosis related carcinoma. On account of rapid development and oc-cult genesis of HCC, most patients were always firstly diagnosed with intrahepatic or ex-trahepatic metastasis at advanced stage, therefore lost the opportunity for optimal surgerytreatment; conversely, the HCC patients who had received timely diagnosis and radicalsurgery at onset stage, could get more survival days. The aforementioned fact shows the early diagnosis of HCC is the most essential factor which determines HCC patients’ sur-vival and prognosis. So far, many molecules are applied to diagnosing HCC in clinic,however, all of them exist the shortage of deficient sensitivity and deficient specificity.Therefore, a novel tumor biomarker for HCC is an urgent need to find.Mature microRNAs (miRNAs), are19-to25-transcript of small non-coding RNAfamily, which play an important role in the processes of physiology and pathophysiology;also, these tiny molecules could act as oncogene or tumor suppressor gene in malignan-cies, regulating the expression of target gene after being transcribed. Several researchesindicate microRNA expression profile has a close relationship with the type of tumor. ThemicroRNA expression is different in diverse tumors, which suggests microRNA could beapplied to the diagnosis or prognosis of neoplasms.In this study, by the means of microRNA microarray, RT-qPCR and statistical analy-sis,4parts had been performed to seek a high-efficient tumor biomarker.Part I: collecting clinical information and selecting enrolled patientsMethod: establishing database using software Epidata3.0; collecting the clinicalinformation of HCC patients and chronic HBV-hepatitis (CH) patients who are meetingthe inclusion criteria; analyzing the comparability between two groups utilizing the soft-ware SPSS21.0.Result: the average serum α-fetoprotein (AFP) value in HCC patient was17.80ng/ml±598.60ng/ml, which was higher than the average value7.10ng/ml±13.10ng/ml in CH patients, there was statistical difference between twogroups (p=0.007). However, no significant difference was observed in other clinical in-formation.Conclusion: serum AFP is a useful biomarker for HCC, and it is increased in mostHCC patients. However, serum AFP has the disadvantages of deficient sensitivity anddeficient specificity. Except serum AFP value, other clinical information between HCCand CH patients are similar, and the two groups are fully comparable.Part II: screening microRNA expression profile of HCC, selecting microRNA-139as objective gene Method: screening microRNA expression profile of HCC using microRNA micro-array; after analyzing the microarray data and reviewing literatures, selecting mi-croRNA-139as objective gene in this study.Result: the result of microRNA microarray was showing the most conspicuous un-der-expression values were found for miRNA-139, miRNA-99a, miRNA-27b, miR-NA-378a, miRNA-378e, and miRNA-30c, while the over-expression values were foundfor miRNA-21, miRNA-221, miRNA-148b, and miRNA-186. After deliberating, we se-lected microRNA-139among the10microRNAs as the objective gene in this study.Conclusion: the microRNA expression profile in this study is lacking conformitywith other studies’results, the possible explaination is the heterogeneities between dif-ferent cases or different specimens, which tells us tumorigenesis is a complex processinvolved with numbers of genes. MicroRNA-139is down-regulated in many neoplasms,however, no microRNA-139related study was reported previously in the aspect of HCCclinical research. So microRNA-139is the feasible objective to be studied in the subse-quent studies.Part III: detecting microRNA-139expression level in tissue and plasma specimensby means of RT-qPCRMethod: extracting total RNA from tissue and plasma using miRNeasy Mini kitand BLOODmisi kit respectively; after confirming the purity of total RNA solution ismeeting the criterion, detecting microRNA-139expression level of each specimen bymeans of RT-qPCRResult: miRNA-139expression level was lower in cancerous tissue and plasma ofHCC patient, compared with peritumoral noncancerous tissue and plasma of CH patientrespectively. Correlation analysis showed the expression quantity of miRNA-139inplasma was positively correlated to that in cancerous tissues.Conclusion: microRNA-139is lowly expressed in hepatocellular carcinoma andother tumors, therefore, we think the down-regulation of plasma miRNA-139expressionmight be a common event in malignancies. Meanwhile, we observed the positive correla-tionship between miRNA-139expression level in tissue specimens and that in plasma specimens, the primary reason for which was explained as that plasma miRNA is releasedfrom tumour-derived microvesicles or exosomes. Due to the defense of microvesicles orexosomes, RNase in blood could not degrade circulating microRNAs.Part IV: the clinical significance of plasma microRNA-139for HCC patientsMethod: based on31HCC patients’clinical information and plasma mi-croRNA-139expression data, analyzing the clinical significance of plasma mi-croRNA-139for HCC patients utilizing software SPSS21.0. There is statistically signif-icant difference while the corresponding p value is smaller than0.05.Result: plasma microRNA-139is a potential diagnostic biomarker for HCC, espe-cially while combined with α-fetoprotein, the sensitivity and specificity went up to90.3%and87.1%respectively. Based on plasma microRNA-139expression level, we divided31HCC patients into two groups, the low group, representing the25patients with plasmamiRNA-139level under-3.240, and the high expression group, representing the remain-ing6ones. The statistical results showed plasma miRNA-139expression level was cor-related with serum AFP value, Edmondson-Steiner grading, DB and CEA value. Moreo-ver, the short-term survival analysis showed the1-year cumulative survival rate of pa-tients in low expression group was48.0%with439survival days (±50days) mediansurvival time, while that in high expression group was66.7%, and the median survivaltime was503days (±69days). Adjusted for Edmondson-Steiner grading, Child-Pughclassification, CLIP scoring and etc, there was statistical difference on the survival ratebetween HCC and CH patients.Conclusion: microRNA-139alone has the high sensitivity up to80%but low speci-ficity when diagnosing HCC, however, whlie combined with serum AFP, the two indexeswas increased observably. In addition, microRNA-139is negatively correlated with Ed-mondson-Steiner grading, which was higher while cancer invasiveness is getting stronger.Finally, the survival analysis was performed. All beforementioned verifies mi-croRNA-139is useful for HCC diagnosis and prognosis, and it might be play a protectiverole in HCC. |
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